Gli3 regulates the proliferation and differentiation of progenitors in the developing neocortex

The cerebral cortex is a highly complex, layered, grey matter structure which plays major roles in processes such as consciousness, memory, and perception. In humans, it is the cortex which bestows us with cognitive abilities unique to our species, in part due to the massive expansion of cortical progenitors during embryonic development. The mature cortex of mammals is composed of a highly heterogenous population of cells, however only the glutamatergic projection neurons are generated within the embryonic neocortex itself. These cells are formed from apical and basal cortical progenitors in the developing dorsal telencephalon, and in the mouse cortical neurogenesis begins at approximately embryonic day 10.5 (E10.5). Regulation of the differentiation of progenitors into neurons is a tightly regulated process, under the control of a strict interplay between signalling molecules and transcription factors. The zinc-finger transcription factor Gli3 plays established roles in regulating the development of the dorsal telencephalon. Through the evaluation of a number of mutants, Gli3 has been shown to function in patterning, the production and fate of cortical neurons, lamination, and axon tract formation, amongst other things. It is shown here that Gli3 is highly expressed in Pax6+ apical progenitors in the dorsal telencephalon, the largest population of progenitors between E11.5 and E12.5. The aim of this thesis was to evaluate how Gli3 regulates the proliferation and differentiation of cortical progenitors into neurons during these time points in the mouse. Embryos of the Gli3Xt/Pdn mutant, which exhibit a reduction in Gli3, were used. At E11.5, the proportions of apical, basal, and early-born cortical neurons were altered in the dorsal telencephalon of the mutant. Microarray analysis revealed an up-regulation of Cdk6 in Gli3Xt/Pdn embryos compared to control. Cdk6 is required for the transition from G1-phase into S-phase during the cell cycle, and shortening of G1-phase has been shown to correlate with progenitors self-renewing instead of differentiating. In agreement with the function of Cdk6, G1-phase and the length of the total cell cycle were shortened in the E11.5 Gli3Xt/Pdn mutant. At E12.5, the length of G1-phase and total cell cycle length were also shortened, and the proportion of apical and basal progenitors and early-born neurons were altered. It appears as though there is a delay in apical progenitor differentiation into basal progenitors and neurons in the Gli3Xt/Pdn embryo, which correlates with the shortening of G1-phase observed. Interestingly, the length of S-phase was also decreased in the mutant at both E11.5 and E12.5, despite no obvious candidates indicative of S-phase regulation being identified in the microarray screen. Taken together, the evidence demonstrates that Gli3 plays a key role in regulating the differentiation of cortical progenitors at the correct time, likely in part due to regulation of the cell cycle, in order to regulate growth of the cerebral cortex

UCP1 expression in human brown adipose tissue is inversely associated with cardiometabolic risk factors.

ObjectiveBrown adipose tissue (BAT) is a therapeutic target for obesity. 18F-Fluorodeoxyglucose positron emission tomography (18F-FDG-PET) is commonly used to quantify human BAT mass and activity. Detectable 18F-FDG uptake by BAT is associated with reduced prevalence of cardiometabolic disease. However, 18F-FDG uptake may not always be a reliable marker of BAT thermogenesis, for example insulin resistance may reduce glucose uptake. Uncoupling protein 1 (UCP1) is the key thermogenic protein in BAT. Therefore, we hypothesized that UCP1 expression may be altered in individuals with cardiometabolic risk factors. MethodsWe quantified UCP1 expression as an alternative marker of thermogenic capacity in BAT and white adipose tissue (WAT) samples (n=53) and in differentiated brown and white pre-adipocytes (n=85). ResultsUCP1 expression in BAT, but not in WAT or brown/white differentiated pre-adipocytes, was reduced with increasing age, obesity and adverse cardiometabolic risk factors such as fasting glucose, insulin and blood pressure. However, UCP1 expression in BAT was preserved in obese subjects of &lt;40 years of age. To determine if BAT activity was also preserved in vivo, we undertook a case-control study, performing 18F-FDG scanning during mild cold exposure in young (mean age ~22y) normal weight and obese volunteers. 18F-FDG uptake by BAT and BAT volume were similar between groups, despite increased insulin resistance. Conclusion18F-FDG uptake by BAT and UCP1 expression are preserved in young obese adults. Older subjects retain precursor cells with the capacity to form new thermogenic adipocytes. These data highlight the therapeutic potential of BAT mass expansion and activation in obesity. <br/

Identifying paediatric nursing-sensitive outcomes in linked administrative health data

There is increasing interest in the contribution of the quality of nursing care to patient outcomes. Due to different casemix and risk profiles, algorithms for administrative health data that identify nursing-sensitive outcomes in adult hospitalised patients may not be applicable to paediatric patients. The study purpose was to test adult algorithms in a paediatric hospital population and make amendments to increase the accuracy of identification of hospital acquitted events. The study also aimed to determine whether the use of linked hospital records improved the likelihood of correctly identifying patient outcomes as nursing sensitive rather than being related to their pre-morbid conditions. Algorithm for nursing-sensitive outcomes used in adult populations have to be amended before application to paediatric populations. Using unlinked individual hospitalisation records to estimate rates of nursing-sensitive outcomes is likely to result in inaccurate rates

Cerebral cortex expression of Gli3 is required for normal development of the lateral olfactory tract

<div><p>Formation of the lateral olfactory tract (LOT) and innervation of the piriform cortex represent fundamental steps to allow the transmission of olfactory information to the cerebral cortex. Several transcription factors, including the zinc finger transcription factor Gli3, influence LOT formation by controlling the development of mitral cells from which LOT axons emanate and/or by specifying the environment through which these axons navigate. <i>Gli3</i> null and hypomorphic mutants display severe defects throughout the territory covered by the developing lateral olfactory tract, making it difficult to identify specific roles for <i>Gli3</i> in its development. Here, we used <i>Emx1Cre</i>;<i>Gli3</i>^<i>fl/fl</i> conditional mutants to investigate LOT formation and colonization of the olfactory cortex in embryos in which loss of <i>Gli3</i> function is restricted to the dorsal telencephalon. These mutants form an olfactory bulb like structure which does not protrude from the telencephalic surface. Nevertheless, mitral cells are formed and their axons enter the piriform cortex though the LOT is shifted medially. Mitral axons also innervate a larger target area consistent with an enlargement of the piriform cortex and form aberrant projections into the deeper layers of the piriform cortex. No obvious differences were found in the expression patterns of key guidance cues. However, we found that an expansion of the piriform cortex temporally coincides with the arrival of LOT axons, suggesting that <i>Gli3</i> affects LOT positioning and target area innervation through controlling the development of the piriform cortex.</p></div

Loss of <i>Dmrt5</i> affects the formation of the subplate and early corticogenesis

Dmrt5 (Dmrta2) and Dmrt3 are key regulators of cortical patterning and progenitor proliferation and differentiation. In this study, we show an altered apical to intermediate progenitor transition, with a delay in SP neurogenesis and premature birth of Ctip(2+) cortical neurons in Dmrt5(-/- )mice. In addition to the cortical progenitors, DMRT5 protein appears present in postmitotic subplate (SP) and marginal zone neurons together with some migrating cortical neurons. We observed the altered split of preplate and the reduced SP and disturbed radial migration of cortical neurons into cortical plate in Dmrt5(-/-) brains and demonstrated an increase in the proportion of multipolar cells in primary neuronal cultures from Dmrt5(-/-)embryonic brains. Dmrt5 affects cortical development with specific time sensitivity that we described in two conditional mice with slightly different deletion time. We only observed a transient SP phenotype at E15.5, but not by E18.5 after early (Dmit5(lox/lox);Emx1(Cre)) but not late (Dmrt5(lox/lox);Nestin(Cre)) deletion of Dmrt5. SP was less disturbed in Dmrt5(lox/lox);Emx1(Cre) and Dmrt3(-/- )brains than in Dmrt5(-/-) and affects dorsomedial cortex more than lateral and caudal cortex. Our study demonstrates a novel function of Dmrt5(-/-) in the regulation of early SP formation and radial cortical neuron migration.Collaborative grant from the Wiener-Anspach Foundation to E.J.B. and Z.M. (Role of the Dmrt5 Transcription Factor in the Development of the Earliest Cortical Circuits); work in the laboratory of E.J.B was supported by grants from the Fund for Scientific Research (FRFC 6973823, CDR 29148846); Walloon Region (First International project "NEURON"); Jean Brachet Foundation; work in the laboratory of Z.M. was funded by Medical Research Council (UK), (G00900901, MR/N026039/1); Royal Society and Anatomical Society. Work in the laboratory of T.T. was supported by the Medical Research Council (MR/K013750/1)

Innervation of the piriform cortex is disorganized in P7 <i>Gli3</i>^<i>cKO</i> brains.

<p><b>(A)</b> In P7 control brains, Satb2 is expressed in layer II/III neocortical neurons and Tbr1 in layer VI neocortical neurons and layer II neurons of the piriform cortex. <b>(B)</b> Magnification of the transition area from neocortex to piriform cortex. <b>(C, D)</b> In <i>Gli3</i>^<i>cKO</i> brains, the transition between neocortex and piriform cortex is shifted medially. <b>(E, F)</b> Ctip2+ neurons are positioned in layer II of the control piriform cortex. <b>(G, H)</b> In <i>Gli3</i>^<i>cKO</i> brains, Ctip2+ neurons occupy a similar layer position but layer II is expanded medially. <b>(I, J)</b> Cellular organization of the piriform cortex visualized by immunofluorescence analysis of CR, Map2 and Dapi. In control brains, CR+ LOT axons extend to layer Ia; Map2 dendrites are present in layer Ia and Ib and Dapi+ cells occupy layer II. <b>(K, L)</b> In <i>Gli3</i>^<i>cKO</i> brains, CR+ LOT axons extend in layer Ia but some CR+ axons aberrantly project in layer Ib and II. Map2+ dendrites are disorganized in layers Ia and Ib. (<b>M</b>, <b>N</b>) <i>Gad67</i> in situ hybridization and Calretinin immunolabeling revealed interneurons and LOT axons in the piriform cortex of control brains, respectively. (<b>O</b>, <b>P</b>) In <i>Gli3</i>^<i>cKO</i> brains, there is no overlap in Gad67 and Calretinin staining in layer II of the piriform cortex. Arrows in A, D, E, H, I and L indicate the position of the rhinal fissure, arrows in N and O indicate <i>Gad67</i>+CR+ interneurons. Scale bars: A-P:250μm.</p

Afferent input from the olfactory bulb expands into more medial positions in the <i>Gli3</i>^<i>cKO</i> cortex.

<p><b>(B, D, H, J)</b> DiI crystal placement in the olfactory bulb (OB) and OB-like structure of control (B, D) and <i>Gli3</i>^<i>cKO</i> brains (H, J), respectively (arrow). (J) In P7 <i>Gli3</i>^<i>cKO</i> brains, the OB protrusion is more prominent compared to E18.5 but not as much as in wild-type brains. <b>(A, C, G, I)</b> Anterograde labeling of LOT axons and their collateral branches in E18.5 (A, C) and P7 (G, I) control brains. In P7 brains (G, I), the LOT occupies the outer piriform cortex layers and DiI labelling extends into layer III. Note the distinct gap between DiI labelling in the LOT and in layer III (I). <b>(E, F)</b> In E18.5 <i>Gli3</i>^<i>cKO</i> brains, the LOT position is shifted medially and the LOT formation appears lense densely packed (arrowhead). <b>(K)</b> A dense population of DiI labelled branches is present in layer III with a barely discernible gap between the LOT and layer III (K, arrowheads). <b>(L)</b> Mitral cell axons occupy a medially expanded region in P7 <i>Gli3</i>^<i>cKO</i> brains. Note the aberrant formation of an axon bundle that projects into the ventral telencephalon (L, <b>arrow</b>). Scale bars: A, G, F and L:50μm; B, D, H and J:0.5μm;C-E, I-K:250μm.</p

Mitral and granule cell layers are formed in the early OB-like primordium of E14.5 <i>Gli3</i>^<i>cKO</i> embryos.

<p><b>(A, B, E, F, I, J)</b> In the olfactory bulb primordium (OBp), Tbr1, <i>Id2</i> and <i>Ap2e</i> are specifically expressed in mitral cells at the rostral tip of the OB of control embryos. <b>(C, D, G, H, K, L)</b> In <i>Gli3</i>^<i>cKO</i> mutants, Tbr1, <i>Id2</i> and <i>Ap2e</i> expressing cells form a thick band at the rostral tip of the OB-like primordium. <b>(M, N)</b> Control embryos show <i>ER81</i> expression in interneuron progenitors in the granule cell layer of the OB primordium. <b>(O-P)</b> In <i>Gli3</i>^<i>cKO</i> mutants, <i>ER81</i> transcripts are present in a distinct cell layer of the OB-like primordium but the <i>ER81</i>+ inner cell layer is extended into the outer mitral cell layer (arrowheads in O). Scale bars: A-P:250μm.</p